1. Field of the Invention
The invention relates to a method for identifying a barley variety using the thermostability of xcex2-amylase encoded by the barley as an indicator. The invention also relates to a method for identifying a barley for brewing using this thermostability of the xcex2-amylase as an indicator. Furthermore, the invention relates to a method of breeding method of a barley variety and a barley variety bred according to this breeding method, using the barley variety identified by the method of the present invention.
2. Description of the Background Art
Malting barley is conventionally bred using a traditional mating technique, for the most part comprising the following two steps. In the first step, individuals having different genotype are mated together and subsequently a selection is conducted from many resultant progeny lines based on desirable cultivation properties such as agricultural properties, adaptability to an environment, and resistance to disease, and a selected line is genetically fixed by growing. This step takes in the neighborhood of 5-6 years. After this long first step, in the second step, a malt analysis of many lines selected and grown in the first step is made, and only good barley lines are selected according to the result of the analysis of their brewing property. The selected barley lines are bred for malting barley.
In conventional barley breeding, however, survey and selection of brewing properties cannot be conducted during the first step. Therefore, a selection and breeding of agronomic character must be performed from grand scale mated progeny lines and then followed by genetic fixation. Moreover, the existence of a good barley line with favorable brewing properties is not guaranteed.
The breeding of a desirable line using this method largely depends on whether an individual breeder can identify a barley having a good brewing property using his experience and observations. Therefore, the breeding of a malting barley having the desired brewing properties must rely on the experience of a brilliant breeder.
Furthermore, the brewing property used as an indicator for selecting a good line in the conventional method largely depends on environmental and climatic conditions. Therefore, the evaluation of characteristics associated with brewing properties must be conducted over the course of several years.
As described above, the conventional method for breeding malting barley with improved brewing properties requires an experienced and skilled barley breeder. Further, even a highly skilled barley breeder requires a long breeding time and tremendous effort to identify malting barley having good brewing properties.
On the other hand, in the past, studies on biochemical differences of xcex2-amylase among barley varieties have been conducted, based on the knowledge that xcex2-amylase is a very important enzyme as a carbohydrase in the brewing of beer. These studies have indicated that there are differences in a gene region coding for xcex2-amylase among barley varieties (Tsuchiya et al. 1995, proceedings of EBC, 109-116). Further, it has been reported that barley varieties can be classified into two types by the isoelectric focusing pattern of xcex2- amylase (Eglinton et al. 1996, Proceedings of V IOC and VII IBGS, 8-10).
However, no relationship between the biological/biochemical properties of xcex2-amylase and the beer brewing property among barley varieties has been found.
As a result of considerable research effort, the inventors have found that there is a relationship between the biological property of xcex2-amylase and the beer brewing property among barley varieties. Using these findings, they have developed a method for identifying varieties that is not affected by natural environment such as climate and the brewing property can be evaluated early after the initiation of breeding.
As described above, the inventors have made numerous research efforts toward the method for identifying a character of interest, by which the breeding period and amount of labor in the breeding of barley is reduced. As a result of this, they have found that there are differences in the degree of a thermostability of xcex2-amylase enzyme activity among barley varieties, and that these differences in thermostability significantly influence the beer brewing property. Further, the research has revealed that the varieties can be identified by differences in thermostability of xcex2-amylase among barley varieties. According to these findings, the inventors have discovered a method for identifying and breeding barley varieties having a good brewing property, or suitability for brewing, using the thermostability of xcex2-amylase as an indicator.
Therefore, the inventive method for identifying barley varieties is characterized by identifying barley varieties, based on the thermostability of xcex2-amylase which the barley varieties encode. Therefore, barley varieties have different thermostability of xcex2-amylase respectively, and it is possible to identify varieties conveniently, based on this thermostability.
The term xe2x80x9cthermostabilityxe2x80x9d as described herein may be expressed as the term xe2x80x9cheat resistancexe2x80x9d which is used to indicate the remaining enzyme activity after heating. Specifically, the thermostability of xcex2-amylase can be expressed, based on the remaining enzyme activity after the heat treatment of, for example 57.5xc2x0 C. for 30 minutes. The treatment conditions are not limited to this temperature and time only but includes any temperature and heating time. Therefore, all such methods are included in the present invention, wherein the thermostability is determined by the remaining enzyme activity after heating and the varieties are identified, based on this thermostability.
Also, as described above, the inventors have discovered that the thermostability of xcex2- amylase significantly influences the brewing property, and have developed a method for identifying a more suitable barley variety for brewing. Therefore, the inventive method for identifying a barley variety allows the selection of a barley variety having a good brewing property by selecting a variety with a greater thermostability of xcex2-amylase.
For example, a suitable variety for brewing is a barley which retains 20% or more, preferably 30% or more, more preferably 40% or more, of the activity of non-heated xcex2- amylase after heating sample such as an extract solution of barley at the above 57.5xc2x0 C. for 30 minutes. This temperature, however, is illustrative only, and the thermostability also can be determined at other than this temperature, and the determination of the thermostability by different temperature and treatment time is included in the present invention.
As described above, unlike the conventional method depending on the experience of a breeder, the present invention makes it possible to objectively identify a variety with a preferable brewing property, based on a physical property of the thermostability of xcex2-amylase of a barley in breeding. This makes it possible to simply and certainly select a barley variety with a preferable brewing property.
Further, the inventors have found that there is a correlation between the thermostability of xcex2-amylase and the isoelectric point value of xcex2-amylase and have also found that the gene coding for xcex2-amylase (the xcex2-amylase gene) has polymorphisms among varieties and that there is a correlation between these polymorphisms and xcex2-amylase. Therefore, these findings allow indirect measurement based on isoelectric points or gene sequences rather than direct measurement of the thermostability of xcex2-amylase which each barley variety possesses. In recent years, advancements in biotechnology have caused the development and improvement of various analytical apparatus and the like, and, by utilizing these, simple measurement based on the isoelectric points or gene sequences, rather than direct measurement of the thermostability, is possible. This in turn makes it possible to more easily identify and select a barley variety with a good brewing property. Of course, these methods for indirectly measuring the thermostability may be employed alone, while it is possible to more effectively identify a desired variety, by combining these methods.
The determination of the above-described thermostability may be conducted using an enzyme solution or nucleic acid solution collected from each barley variety. More particularly, when determining the thermostability after heating, or when determining the isoelectric point, the enzyme solution collected from seeds or the like of each barley variety can be used. These seeds for obtaining the enzyme solution can be used in any growth stage, but fully ripe seeds are preferably used. Further, when determining polymorphisms of xcex2- amylase, the nucleic acid solution collected from tissues and cells of, for example, green leaves of each barley variety can be utilized. Any growth stage of tissues and cells can be used herein, regardless of their growth stage. Therefore, by obtaining the enzyme solution or nucleic acid solution from an early growth stage of barley, it is possible to dramatically shorten the time required to judge as compared to a conventional method, and it is possible to rapidly identify a variety with a brewing property. This allows not only an advantage with respect to time-saving but also reduces of the cost of examination associated with the conventional method.
Further, a barley variety having the desired thermostability of xcex2-amylase, or a barley variety with a good brewing property or greater suitability for brewing can be bred according to the present invention. As a result of this, it is possible to maintain, improve, or develop the quality of a beer product by utilizing the barley variety bred herein for manufacturing beer.
Moreover, it is possible to provide a better brewed product by using the barley variety having such good brewing property for manufacturing the brewed product.
A more complete appreciation of the present invention and many of the attendant advantages thereof will be readily obtained as the same becomes better understood by reference to the following detailed description when considered in connection with the following drawings.